Expression Of Fas/FasL In Rat Spinal Cord Apoptosis Induced By Ropivacaine Hydrochloride

Objective: Spinal canal anesthesia can induce severe neurological complications,which may be related to the neurotoxic effects of local anesthetics on the spinal cord.At present,the specific mechanism of neurotoxicity of local anesthetics is still inconclusive,and the induction of neuronal apoptosis may be an important factor.Fas / FasL is an important way for cells to exogenous apoptosis.Studies have shown that Fas / FasL plays an important role in neuronal apoptosis caused by acute spinal cord injury,but its role in ropivacaine-induced spinal cord injury is still unknown.The purpose of this study was to investigate whether the exogenous apoptosis pathway mediated by Fas / FasL is involved in the process of spinal cord injury and apoptosis induced by ropivacaine hydrochloride.Methods: Twenty-four male SD rats were randomly divided into 4 groups and made into the intrathecal catheterization model.After 3 days of postoperative observation,the rats with well modeling were treated differently according to the group.The normal saline group: 0.12 ml / kg of intrathecal injection of normal saline;0.5% group: intrathecal injection of 0.5 % Ropivacaine;1% group: intrathecal injection of the same dose of 1% ropivacaine;2% group: intrathecal injection of the same dose of 2% ropivacaine.Each group of drugs was injected intrathecally 8 times,with an interval of 1.5 hours.Changes in rat Paw Mechanical Retraction Threshold(PMWT)were measured before the intrathecal catheterization,before intrathecal administration,and 24 hours after intrathecal administration.After completing all PMWT tests,the rats were sacrificed and the lumbar swelling of the spinal cord was collected.Hematoxylin-eosin(HE)staining was used to observe the morphological changes in spinal cord.TUNEL staining was used to detect the apoptosis of spinal cord cells in each group,Quantitative Real-time PCR(qPCR)was used to detect the changes of Fas,FasL,caspase-3 and caspase-8 m RNA levels in each group.Immunohistochemical staining and Western Blotting were used to detect Changes of Fas,FasL,caspase-3 and cleaved caspase-3 cleaved caspase-3 in protein levels in each spinal cord.Results: There was no significant difference in PMWT before and after the intrathecal catheterization were found in each group.Compared with those before intrathecal administration,the PMWT values in the 1% group and the 2% group significantly increased at 24 hours after administration(P <0.05).HE staining showed that the spinal cord tissue in the saline group was intact and dense,and the cell morphology was clear and good.The degree of spinal interstitial edema in each of the ropivacaine-treated groups increased with the increase of drug concentration,and some of the cell nucleus contracted and disappeared in the 1% and 2% groups.TUNEL staining showed that only a small amount of green fluorescence was scattered in the normal saline group,and there were fewer apoptotic cells.While in each ropivacaine group,as the drug concentration increased,apoptotic cells gradually increased.The dorsal horn apoptotic rate in the saline group was lower than that in all ropivacaine-treated groups under high magnification(all P <0.05).Compared with the 0.5% group,the apoptosis rate of spinal cord cells was higher in the 1% and 2% groups,and the difference was statistically significant(P <0.05).The qPCR results showed that compared with the normal saline group,the m RNA expression of Fas,FasL,caspase-3,and caspase-8 in each ropivacaine-treated group was up-regulated with increasing drug concentration,and the difference was statistically significant(P <0.05)Immunohistochemistry and Western Blotting results showed that the expression of Fas,FasL,caspase-3,and cleaved caspase-3 in spinal cord tissues increased with drug concentration compared with the saline group(P <0.05).Conclusion: Ropivacaine hydrochloride can cause PMWT value changes in rats,and can induce pathological damage to the spinal cord of rats with increased apoptotic cells.The mechanism of ropivacaine-induced apoptosis in rat spinal cord cells may be related to up-regulation of Fas / FasL.